Electrochemical sensor

ABSTRACT

The invention relates to a sensor for detecting substance concentration or activity or for determining the presence of substances based on electrochemical reactions. The electrochemical sensor comprises an electrode with surfaces inside the electrode, wherein electrochemical detection reactions occur. Said sensor guarantees high flow densities and is suitable for miniaturization. A substance-recognizing agent is advantageously placed in the electrode in contact with the inner surfaces. This enables not only short response times and long service life but also measurements with excellent linearity.

The invention relates to a sensor for determining materialconcentrations, activities or for material detection on the basis ofelectrochemical reactions.

A structure often used in electrochemical sensors consists of a flatsupport, on which is arranged a flat detection electrode. This electrodeis in contact with the sample medium. Certain materials, in the case ofplatinum electrodes for example hydrogen peroxide, may be detectedelectrochemically as a result of electrochemical reactions on theelectrode surface.

The disadvantage of sensor arrangements of this type is thecomparatively small electrode surface and the often low currentdensities resulting therefrom. Particularly in the course of advancingminiaturization, the surface dimensions of flat electrodes required toachieve sufficiently high current densities are often an insurmountablelimit to smaller sensor structural shapes.

The flat sensor electrode is often in contact with amaterial-recognizing substance in the form of a thin membrane. Materialswhich may be detected electrochemically at the electrode are formed inthe membrane due to a specific detection reaction.

A thin membrane guarantees short diffusion paths from the site of thechemical detection reaction to the electrode and hence short responsetimes. A further advantage of thin membranes is also the avoidance ofsubstrate limitations. In the case of glucose sensors based on glucoseoxidase by way of example, there is the danger of oxygen limitation andhence of undesirable non-linearities if the oxygen required for thedetection reaction cannot diffuse into the membrane in sufficientquantity.

However, these preferences for thin material-recognizing membranes arepartly compensated by adhesion problems due to the often difficultbinding of the thin membrane to support and electrode as well as byencapsulation problems. A further disadvantage of thin membranes is theshort service life of such sensors, since the comparatively few activemembrane components due to the low membrane volume are deactivated orspent in a short time.

The disadvantages just mentioned can be partly avoided by using thickermaterial-recognizing membranes. However, for thicker membranes there isthe problem that long response times for the sensor have to be takeninto account due to the long diffusion paths from the site of thechemical reaction to the flat electrode arranged on the support.Furthermore, some of the reacted materials may diffuse from thematerial-recognizing membrane before detection at the electrode surface,as a result of which the sensitivity is reduced.

A problem with sensor cross-sensitivities often occurs independently ofthe membrane thickness. The required minimization of suchcross-sensitivities is often associated with complex and thereforeexpensive modifications of conventional sensors.

The object of the invention is to overcome the disadvantages of thestate of the art and to provide an electrochemical sensor which can beminiaturized to analyze liquid or gaseous samples, which has shortresponse times and at the same time an increased linear measuring rangeas well as a long service life, particularly in combination withmaterial-recognizing substances.

For an electrochemical sensor which has at least one electrode havinginner hollow cavities into which the material to be determined and/orreacted reaction products may enter, and in which a material-recognizingsubstance is incorporated in these cavities at least in some regions,advantageously large active detection surfaces can be achieved in theelectrode interior even for small external electrode dimensions. Asensor of this type permits realization of high current densities and isparticularly suitable also for miniaturization.

Electrodes having inner hollow cavities and consequently having innersurfaces have, in contrast to the traditional flat electrodes, anessentially three-dimensional functionality. Electrodes of this type mayhave, for example a latticed, reticulated, filamentary or porousstructure. It is important that pores, tubes or other hollow cavitieshaving surfaces are present in the interior of the electrode, by meansof which the medium to be analyzed and the material to be detected orreacted reaction products of the material to be detected may come intocontact with the electrode surface.

The advantages of sensors based on thin-layer membranes and of thosebased on thick-layer membranes can be combined in a sensor havingmaterial-recognizing substance incorporated in the sensor electrode.Consistently short response times are achieved by incorporating thematerial-recognizing substance in the electrodes, in contrast tothick-layer membrane sensors, independently of the volume of thematerial-recognizing substances. This is due to the fact that thediffusion path between the site of the chemical reaction and the site ofdetection of the reaction on the electrode surface is minimal everywherein the electrode interior. In addition to short response times, highsensitivities can be achieved for amperometric sensors. Since the volumeof material-recognizing substance may be increased without considerablelosses regarding the response times, a high number of active componentsare available. The operating period of the sensor is thus considerablyextended.

For sensors having material-recognizing substance incorporated in theelectrode, it may be advantageous to increase the hollow cavity volumesin favor of incorporating a larger quantity of material-recognizingsubstance for constant external electrode dimensions. Optimization ofsensor service life and sensitivity adapted to the particularapplication is possible in this manner with consistently short responsetimes.

The essentially three-dimensional sensor electrode is preferably incontact with an electric leakage arranaed on the support and isadvantageously composed of several part electrodes (multi-layerstructure). In individual or in all part electrodes a specificmaterial-recognizing substance is introduced at least in some areas, forexample by capillary forces. Hence the same or differentmaterial-recognizing substances may be embedded in different partelectrodes. Embedding can often be carried out more simply in amulti-layer structure than for solid electrodes having inner surfaces. Alayer electrode which permits simple, layer-like production ofindividual part electrodes is produced in this manner.

Individual part electrodes may be in electrical contact with one anotheron the surface so that only one single part matrix needs to be connectedto an electric leakage. However, layers of an electrically insulatingmaterial may also be arranged between individual part electrodespreferably provided with separate leakages and may be permeable in eachcase to the material to be detected and to reaction-assisting orreaction-accompanying materials. The part electrodes may also beseparated from one another by distancing layers (spacer) which haveperforations in a central electrode region. The sandwich constructionpermits incorporation of different material-recognizing substances indifferent part electrodes and simultaneously detection of a plurality ofmaterials in a sample solution.

The electrode or part electrodes may consist of a conductive parentsubstance, for example of metal, or of a conductively coated ormetallised, non-conductive parent substance. Suitable conductive parentsubstances are, for example of metals, such as platinum, silver or goldor of a paste containing carbon and/or one of the afore-mentionedmetals. These materials are also suitable as conductive coating fornon-conductive parent substances.

Metallic parent substances having inner surface can be produced, forexample by etching or by laser treatment. Non-conductive parentsubstances on the other hand often already have an inner surface fromthe start. Papers, such as filter paper, paperboards, glass fibers,plastic fibers, textiles, ceramics, mineral materials or materials ofvegetable or animal origin, are suitable as non-conductive parentsubstances for electrodes or part electrodes. Application ofmetallisation may be effected, for example by sputtering, vaporizationusing pastes or adhesives or by chemical reaction.

The three-dimensional electrode preferably consists of an insulatingparent substance which is surrounded as completely as possible by a verythin layer of a conductor. The production costs of the sensor can beconsiderably reduced by minimizing the metallisation thickness.Furthermore, it is possible to provide larger surface areas of theparent substance (for example paper webs) with metallisationsimultaneously and uniformly. Smaller pieces of it are then used as anelectrode or part electrode for the sensors. The production costs canthus be reduced and the signal reproducibility increased.

The material-recognizing substance incorporated in the sensor electrodepreferably contains at least one active component, such as enzymes,microbes, bacteria or yeasts, which is preferably immobilized using atleast one gel-like or elasticized polymer, such as polyvinyl alcohol,polyvinyl chloride, polyurethane, acrylate or silicone. Thematerial-recognizing substance may be introduced into the electrodeparent substance by utilizing capillary action, by vacuum infiltrationor by pressure filling.

One or more three-dimensional support cavities receiving the at leastone electrode are advantageously arranged on the sensor support or inthe support or at least partly in the support of the electrochemicalsensor. This at least one support cavity may advantageously be providedwith at least one cover. The electrode may thus be encapsulated toprovide protection in simple manner and therefore cost-effectively andfixed on the support. This method of encapsulation also avoids theproblem of membrane adhesion which often occurs in sensors based onthin-layer membranes.

The support and/or at least one of the covers advantageously has anopening which permits entry of the substance to be detected into thesupport cavity. The sensor behavior can be optimized by suitableselection of the opening diameter. A small opening diameter, for examplereduces the outward diffusion of the substance to be detected and thereacted substance.

Support and cover advantageously consist of a film material based on,for example polyethylene, polyester, polyvinyl chloride, polypropylene,polytetrafluoroethylene, cellulose acetate, silicone or a combination ofthese materials. The material should be permeable to reaction-assistingor reaction-accompanying materials, but impermeable to the material tobe determined. The linear measuring range of the electrochemical sensoris extended in this manner, since depletion of the reaction-assisting orreaction-accompanying materials, that is a substrate limitation, isdelayed. Particularly well suited material for support and cover arefilms, such as heat-sealing films, laminating films, self-adhesive filmsor sealable films which are joined to one another by melting oradhesion.

An electric leakage arranged on the support and contacting the electrodepreferably consists of a metal, such as platinum, silver or gold or of apaste containing carbon and/or one of the afore-mentioned metals, andmay be applied to the support by a screen printing process, asilk-screen printing process, a dispensing process, a spraying process,by vaporization or by sputtering.

In addition, a reference electrode or counter-electrode may be arrangedon the support to have a reference which is not influenced by the samplematrix or the reaction products, and to facilitate current flow forvoltametric measurements. A silver/silver chloride paste by way ofexample is a suitable material for this electrode.

Measuring electrode as well as reference electrode or counter-electrodemay come into contact with the medium to be analyzed, for example via aflow channel arranged on the support. A sensor having integrated flowchannel is particularly suited for automation of the detection processor for long-term monitoring. However, direct contact with the samplemedium can also be conceived by immersion or trickling.

The structure of the electrochemical sensor of the invention permitsminimizing of the influence of interfering materials and hencecross-sensitivities in simple manner. This is possible in two ways.Firstly, an interference protective layer may be arranged between sampleto be analyzed and the electrode which prevents entry of interferingmaterials into the support cavity by size and/or charge exclusion.Layers of this type may consist, for example of polymer layers ofsilicone, polytetrafluoroethylene, polyacrylate, polyurethane, celluloseacetate, Naflon, polycarbonate or polyvinyl chloride. They may line thesupport cavity completely or even partly, for example in the region ofthe support or cover opening.

A further possibility for reducing the effect of interfering materialsconsists in arranging a further electrode, preferably in the region ofthe support or cover opening, between the detection electrode and thesample to be analyzed and connecting to a voltage source. Interferingmaterials may be oxidized or reduced by applying a voltage to thiselectrode, resulting in it being possible to reduce cross-sensitivities.The further electrode is advantageously situated in the immediatevicinity of the detection electrode having inner surfaces and iselectrically separated from the latter by an insulating layer or a thingap.

The further electrode may be designed like the detection electrode, butwherein material-recognizing substance is not incorporated in the hollowcavities. This guarantees that interfering materials are reduced oroxidized at the surface of the further electrode on their path throughthe further electrode to the detection electrode with considerableprobability.

Further positive properties and preferences of the invention can be seenfrom the exemplary embodiments described below and the figures.

BRIEF DESCRIPTION OF THE DRAWINGS

FIGS. 1, 2 show an electrochemical sensor (plan view and section A-A′),

FIGS. 3, 4, 5 show an electrochemical sensor with integrated referenceelectrode (plan view, sections A-A′ and B-B′),

FIGS. 6, 7 show an electrochemical sensor with individual contact of thepart electrodes (plan view and section A-A′),

FIGS. 8, 9 show an electrochemical sensor with integrated referenceelectrode in through-flow arrangement (plan view and section A-A′),

FIGS. 10, 11 show an electrochemical sensor with interference protectivelayer (plan view and section A-A′),

FIGS. 12, 13 show an electrochemical sensor with particularly highco-substrate permeability (plan view and section A-A′), and

FIG. 14 shows the calibration curve of an electrochemical sensor of theinvention having increased co-substrate permeability.

Glucose can be determined by way of example using the electrochemicalsensor shown in FIG. 1 and FIG. 2. In this case a 250 μm thicklaminating film made from polyethylene/polyester is used for the support1. The leakage 2 consists of a carbon paste deposited on the support 1by means of screen printing and extends into the electrode cavity 4. Ametallised electrode made of three thin filter papers 7 of 3 mm diametersputtered with platinum is situated in the electrode cavity 4. Theindividual layers are in surface electrical contact and in theirentirety form the three-dimensional, irregularly reticulated electrode.The cover 3 consists of a 60 μm thick laminating film made frompolyethylene/polyester, which is permeable to oxygen over the entiresurface area, whereas it is permeable to glucose only through thepreviously punched opening 5 having a diameter of 1 mm. The diffusion ofglucose into the support cavity 4 is possible via this opening. Thematerial-recognizing membrane consisting of a solution of glucoseoxidase in a photo-crosslinkable polyvinyl alcohol gel, is introducedinto the pores of the metallised electrode cavity 4 through the opening5. Distribution of the membrane in the support cavity 4 is thus effectedby capillary forces. The material-recognizing membrane is polymerized byUV radiation and thus rendered water-insoluble. The support cavity 4 isa reservoir for the material-recognizing membrane and thus extends theservice life of the sensor.

FIG. 14 shows the calibration curve of a sensor of this type. The goodlinearity, even for high glucose concentrations up to 35 mmole/liter,due to the improved co-substrate permeability can be seen clearly. Fortraditional sensors the end of the linear range lies at considerablylower glucose concentrations.

Determination of a number of other substances is possible using anelectrochemical sensor of this type, in addition to glucosedetermination, wherein essentially the composition should be adapted tothe material-recognizing membrane. Some examples are outlined in thefollowing table:

Material- recognizing Analyte substance Co-substrate DetectionCholesterol Cholesterol Oxygen Hydrogen oxidase peroxide Triglycer-Esterase, Oxygen Hydrogen ides glycero- peroxide kinase, glycero-phosphat- oxidase Creatinine Creatinin- Oxygen Hydrogen ase, peroxidecreatinase, sacrosin- oxidase Uric acid Uricase Oxygen Hydrogen peroxide

Integration of the counter-electrode 8 is shown in FIGS. 3, 4 and 5. Ina double-electrode arrangement, the counter-electrode 8 also assumes thefunction of the reference electrode. A silver/silver chloride paste,which is applied to the support 1 by means of screen printing, serves asmaterial for this.

Contact of individual support cavities 4 and 4′ by means of separateleakages 2 and 2′ is shown as the next example of the invention in FIGS.6 and 7. The leakages 2 and 2′ are separated from one another by aninsulating layer (spacer) 6.

The separated support cavities 4 and 4′ are joined to one another by theopening 5′ in the insulating layer 6. Two materials can be detected atthe same time using this sensor configuration by introducing differentmaterial-recognizing membranes into the support cavities 4 and 4′.

In addition, there is the possibility of only filling the electrode insupport cavity 4 with a material-recognizing membrane. For the exampleof glucose determination, the electrode incorporated in the supportcavity 4′ and connected to a voltage source then serves to oxidizeoxidisable substances, such as ascorbic acid or uric acid, at 700 mVagainst a silver/silver chloride paste, so that they no longer interferein the determination in the support cavity 4.

A further modification of the electrode is shown in FIGS. 8 and 9. Aflow channel 9 is inserted here between the electrode cavity 4 or thereference electrode 8 and the cover 3, as a result of which adaptationof the sensor for through-flow measurements is facilitated withoutconsiderable expense.

FIGS. 10 and 11 shows a further electrode of the invention, in which aninterference protective layer is applied above the support cavity 4.

FIGS. 12 and 13 shows a further example of the sensor of the invention.A larger hole as the opening 5 of the support cavity 4 is punched in thecover 3 made from laminating film. The support cavity 4 is covered by alayer 11, which has a particularly high co-substrate permeability and inwhich the opening 5 is introduced. 30 μm thin silicone film, a materialhaving very high oxygen permeability, is used here for the glucosesensor. Materials which cannot be melted or adhered particularly easilycan also be used for the sensor structure in this manner.

What is claimed is:
 1. Electrochemical sensor for determining materialconcentration comprising a support and an electrode arranged in a regionof the support, wherein the electrode comprises a plurality of electrodelayers arranged one above another, wherein two of the plurality ofelectrode layers each comprises a non-conductive parent substance,wherein the non-conductive parent substance of at least one electrodelayer has inner hollow cavities within it, wherein at least some of asurface of the inner hollow cavities within the parent substance iscoated with a layer of a conductor, and wherein a specificmaterial-recognizing substance is incorporated into at least some of theinner hollow cavities within the non-conductive parent substance of atleast one electrode layer.
 2. Electrochemical sensor according to claim1 wherein a spacer comprising an electrically insulating material isarranged between the electrode and another electrode.
 3. Electrochemicalsensor according to claim 2 wherein the two electrodes are provided withseparate leakages.
 4. Electrochemical sensor according to claim 1wherein the non-conductive parent substance comprises an elementselected from the group consisting of: paper, filter paper, paperboard,glass fibers, plastic fibers, textile, ceramic, mineral material, andmaterial of vegetable or animal origin.
 5. Electrochemical sensoraccording to claim 1 wherein the material-recognizing substancecomprises an element selected from the group consisting of: enzymes,microbes, bacteria, and yeasts.
 6. Electrochemical sensor according toclaim 1 wherein the material-recognizing substance is immobilized withthe aid of an element selected from the group consisting of: gelpolymer, elasticized polymer, polyvinyl alcohol, polyvinyl chloride,polyurethane, acrylate, and silicone.
 7. Electrochemical sensoraccording to claim 1 wherein the support comprises a cavity receiving atleast one electrode layer.
 8. Electrochemical sensor according to claim7 wherein the support cavity is provided with a cover. 9.Electrochemical sensor according to claim 8 wherein the support and/orthe cover is permeable to reaction-assisting or accompanying materialsand is impermeable to a particular material to be determined. 10.Electrochemical sensor according to claim 8 wherein the support and/orthe cover has at least one opening.
 11. Electrochemical sensor accordingto claim 8 wherein the support and/or the cover comprises an elementselected from the group consisting of: beat-sealing film, laminatingfilm, self-adhesive film, and sealable film.
 12. Electrochemical sensoraccording to claim 1, wherein a reference electrode and/orcounter-electrode is arranged on the support.
 13. Electrochemical sensoraccording to claim 12 wherein the electrode and the reference electrodeand/or counter-electrode come into contact with a sample medium to beanalyzed via a flow channel.
 14. Electrochemical sensor according toclaim 1 wherein an interference protective layer is arranged between asample medium to be analyzed and an electrode layer.
 15. Electrochemicalsensor according to claim 14 wherein the interference protective layercomprises an element selected from the group consisting of: polymerlayer of silicone, polytetrafluoroethylene, polyacrylate, polyurethane,cellulose acetate, perfluorinated membrane, polycarbonate, and polyvinylchloride.
 16. Electrochemical sensor according to claim 1 furthercomprising another electrode connected to a voltage source and arrangedbetween a sample medium to be analyzed and at least one electrode layer.17. Electrochemical sensor according to claim 16 wherein the supportcomprises a cavity receiving at least one electrode layer, wherein thesupport cavity is provided with a cover, wherein the cover has at leastone opening, and wherein the received electrode layer is arranged in aregion of the opening of the cover.
 18. Electrochemical sensor accordingto claim 1 wherein the support comprises a cavity receiving at least oneelectrode layer, wherein the support cavity is provided with a cover,wherein the material-recognizing substance contains glucose oxidase fordetermining glucose, and wherein the support or the cover is permeableto oxygen.
 19. Electrochemical sensor according to claim 1 wherein theconductor comprises an element selected from the group consisting of:platinum, silver, gold, and carbon.